The proposed studies are directed at understanding how single nucleotide polymorphisms, insertion/deletions, or other mutational changes contribute to the emergence of S. enterica serovars adapted to warm-blooded hosts (including domesticated animals) and which cause typhoid or non-typhoid infections in humans. Our aim is to examine every gene shared by specific pathogenic lineages of S. enterica as being a potential target for positive selection for mutations of a pathoadaptive nature. To do this, we will employ comparative genetic analyses of Salmonella isolates adapted to cold- and warm-blooded hosts and, among the latter, those causing systemic, non-systemic and asymptomatic infections. We expect to use 30-40 fully sequenced genomes for the analysis and about the same or a larger number of strain pairs will be resequenced by mutation tiling microarrays. Genetic polymorphisms will be analyzed for a footprint of positive selection. Using additional isolates, we will investigate association of the mutational changes with strains from specific habitats or types of infection caused. The pathoadaptive nature of mutations in some of the genetic loci will be examined experimentally, using both in vitro and in vivo models. As a result, we plan to obtain (i) a map of S. enterica genes targeted for mutations that contribute to the host adaptation and virulence of S. enterica;(ii) a list of naturally-occurring mutations in these genes that likely affect the gene/protein function in a pathoadaptive manner, and (iii) a comprehensive phylogenetic history and dynamics model of the evolution of virulence in Salmonella. Narrative. We propose to dissect molecular basis of evolution of virulence of Salmonella strains that cause human infections. We will determine genetic changes that contribute to emergence of Salmonella that are infecting warm-blooded animals and cause typhoid or non-typhoid disease in humans.